Effect of Cytoskeletal Disruption on Mechanotransduction of Hydrostatic Pressure by C3H10T1/2 Murine Fibroblasts.
Effect of Cytoskeletal Disruption on Mechanotransduction of Hydrostatic Pressure by C3H10T1/2 Murine Fibroblasts.
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DOI：
10.2174/1874325000802010155
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发表时间：
2008-12-29
期刊：
影响因子：
--
通讯作者：
Elder SH
中科院分区：
文献类型：
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作者：
Shim JW;Wise DA;Elder SH
生理量级（< 10 MPa）的循环静水压力刺激间充质干细胞的软骨分化，但机械转导机制尚未完全了解。据推测，一个完整的细胞骨架将需要不受抑制的机械转导的静水压力。因此，我们研究了选择性干扰肌动蛋白和微管蛋白聚合的药物对压力诱导的聚集蛋白聚糖和col 2a 1（II型胶原）mRNA表达上调的影响。将C3 H10 T1/2细胞在4μM细胞松弛素D或4μM诺考达唑中培养为沉淀，并进行3天的循环静水压力（1 Hz，5 MPa，每天2 h）。鬼笔环肽染色和抗α-微管蛋白抗体的间接免疫染色分别证实了微丝和微管组装的破坏。真实的时间RT-PCR显示，这两种药物基本上降低了聚集蛋白聚糖和col 2a 1 mRNA的基础水平，但没有一种药物阻止相对于改变的基础状态的压力刺激的基因表达增加。因此，上调大分子基因表达的循环静水压力并不需要一个完整的细胞骨架。
Cyclic hydrostatic pressure of physiological magnitude (< 10 MPa) stimulates chondrogenic differentiation of mesenchymal stem cells, but mechanotransduction mechanisms are not well understood. It was hypothesized that an intact cytoskeleton would be required for uninhibited mechanotransduction of hydrostatic pressure. Therefore we examined the effects of drugs which selectively interfere with actin and tubulin polymerization on pressure-induced upregulation of aggrecan and col2a1 (type II collagen) mRNA expression. C3H10T1/2 cells were cultured as pellets in either 4µM cytochalasin D or 4µM nocodazole and subjected to 3 days of cyclic hydrostatic compression (1 Hz, 5 MPa, 2 h per day). Phalloidin staining and indirect immunostaining with anti α-tubulin antibody confirmed disruption of microfilament and microtubule assemblies, respectively. Real time RT-PCR revealed that both drugs substantially lowered the basal level of aggrecan and col2a1 mRNA, but that neither drug prevented a pressure-stimulated increase in gene expression relative to the altered basal state. Thus upregulation of macromolecular gene expression by cyclic hydrostatic pressure did not require a completely intact cytoskeleton.